Background

Bacterial bioinoculate is one of environmentally friendly input for crop production. Several works revealed that effectiveness of Rhizobium bioinoculant have been affected by the native rhizobia population. Hence, this study was initiated to evaluate the effect of selected Bradyrhizobium in nodulating peanut in the major growing areas of Ethiopia.

Methods

The effect of eight effective Bradyrhziobium vis-a-vis 20 kg N ha−1 and the control check on the nodulation and yield of two peanut cultivars (Sedi and Werer 942) at Babile and Fedis, eastern Ethiopia, during 2013 cropping season was evaluated.

Results

The results showed that the inoculation, location and cultivar and their two and three ways interaction had a significant influence on the nodulation and grain yield. Inoculation of Bradyrhizobium increased the nodule number, nodule dry weight, number of seeds per plants, total pods weight, hundred seeds weight, total biomass yield, grain yield and shelling % up to 37, 24.9, 22.4, 22.5, 5.5, 26.2, 35.8 and 24.1%, respectively. Isolate HUGNR-29 was found to be more effective as it scored the highest nodule dry weight and grain yield. Application of inorganic N increased the yield and yield components of peanut in either of locations. Though the statistically better effective nodule % was recorded at Babile, significantly higher nodule number, nodule dry weight, and total N accumulation were obtained from Fedis than Babile site. Significantly higher total pods weight, hundred seeds weight, total biomass weight, grain yield and total plant N accumulation was recorded with the cultivar werer-962 than those produced with Bati-Sedi.

Conclusions

The result showed that Bradyrhizobium inoculation is essential to improve the peanut production through environmentally and economically sustainable ways though the soil native rhizobia nodulating peanut was high. Peanut cultivar and study site based rhizobia inoculant development is also important.

The peanut is the fourth most widely cultivated oilseed in the America continents, Africa and Asia. Peanut plants grow best in well-drained sandy soils and sunny warm temperatures with moderate rainfall. In Ethiopia, the total planted area was 443000 ha and a production of 103.7 M ton of grain as it has been reported in the 2014 cropping season (FAOSTAT 2014). This report also showed that the productivity of peanut has been 1604.1 kg ha−1. To increase this productivity, plant breeder has tried to release new high yielder varieties of peanut in Ethiopia (Kebede and Bushra 2012). So far, peanut varieties improving program in Ethiopian has obtained the peanut variety yielding up to 2.2 ton ha−1. When its productivity is comparing to the productivity (3.25 ton ha−1) that has been reported in North Carolina is very low (Jones 2003). Hence, improving the production and productivity of peanut in Ethiopia using environmental friendly technologies is essential.

Peanut is considered a highly promiscuous species because it is nodulated by rhizobia able to nodulate a diverse group of legumes and a large group of unrelated rhizobia (Alwi et al. 1989). For instance, up to 18% of the tested fast and slow growing rhizobia isolated from cowpea are able to form effective nodules with peanut (Thies et al. 1991; Mpepereki et al. 1996). Several studies have also already shown phenotypically and genetically diverse bradyrhizobia isolates nodulating peanut (Li et al. 1999; Zhang et al. 1999). Most commonly reported isolates of bradyrhizobia nodulating peanut are Bradyrhizobium japonicum and Bradyrhizobium elkanii (Willems et al. 2001; Yang et al. 2005; Nievas et al. 2012; Bogino et al. 2010). Peanut is also nodulated by fast-growing rhizobia such as Rhizobium giardini and Rhizobium tropici (Huang 1990; Taurian et al. 2006). The contribution of biological nitrogen fixation (BNF) through peanut-rhizobia symbiosis has been reported as 40.9 kg ha−1 (Okito et al. 2004).

The presence of nodules on roots of the peanut plant does not necessarily mean that sufficient N2 is being fixed for maximum growth of the host plant (Weaver 1974; Nambiar et al. 1982a). Inoculation is a major strategy for the development of sustainable input of nitrogen into agricultural soils (Lindström et al. 2010). Inoculation of legume seed is an efficient and convenient way of introducing highly efficient rhizobia to the soil to improve the peanut production (Deaker et al. 2004). Inoculation of Bradyrhizobium peanut resulted in the nitrogen fixation amounting up to 186 kg N ha−1 (Moawad et al. 1986). Castro et al. (1999) found that peanut roots have been well nodulated by native strain rather than the exotic one. Hence, selection of efficient strains based on their local ecologic adaptation can increase grain production of crops (Denton et al. 2003; Lindström et al. 2010; Bianco et al. 2013). On top of differing the rhizobia in their ability to fix N2, the effective peanut-rhizobia symbiosis is the product of rhizobial and plant cultivars as well as of environmental conditions (Nelson 1987; Sessitsch et al. 2002). Manipulating the peanut cultivar and Bradyrhizobium strain together in one system would be the most important for a productive approach to optimizing N2 fixation and increase the peanut productivity (Chen et al. 2003). Hence, the hypothesis of this work was stated that inoculation of rhizobia isolated from native soil improves the nodulation and yield of peanut even though the soil native rhizobia population has been > 102 g−1 soil. Our approach has been to utilize indigenous rather than exotic rhizobia, since the former, already adapted to the existing ecological conditions, can provide very satisfactory results (Douka and Xenoulis 1987; Hungria et al. 1996; Makatiani and Odee 2007). Therefore, the objective this work was to evaluate the symbiotic effectiveness of selected indigenous Bradyrhizobium sp. nodulating peanut in eastern Ethiopia.

Description of the study sites

Field studies were conducted in 2013 cropping season at Fedis (09°06.941′N and 042°04.835′E at an altitude of 5476 ft above sea level [asl]) and Babile (09°13.234′N and 042°19.407′E at 5478 ft asl) Agricultural Research Sub-center, Haramaya University, located in the eastern part of Ethiopia. The soils of the study sites (Babile and Fedis) were sandy loam and silty clay loam, respectively. Fedis site receives a mean annual rainfall of 714.3 mm (Ethiopia Meteorological Report, unpublished), with a mean maximum and minimum temperature of 27.8 and 8.8 °C, respectively. The area has a bi-modal pattern, falling in two seasons, with the short rain season occurring between March and May and the long rain season from between June through October. At Babile site, the annual means maximum and minimum temperatures are 20 and 90 °C, respectively with annual mean rainfall of 540 mm which has been calculated based on the weather data between 2000 and 2014. The soil of Babile site before planting the experiment showed the following properties: pH 6.66; EC, 0.04 dS m−1; organic carbon content, 0.56%; total N, 0.06% available P content (Olsen), 2.22 mg kg−1; contents of exchangeable K1+, Ca2+, Mg2+ and Na1+, 0.34, 4.18, 3.5 and 0.15 cmol(+) kg−1, respectively; cation exchange capacity, 8.17 cmol(+) kg−1, number of rhizobia, 5.8 × 102 cells g−l of dry soil. The top 20 cm of Fedis soil contained 1.32% soil organic C, 0.12% total N, 1.78 mg kg−1 Olsen-P, cation exchange capacity, 37.2 cmol(+) kg−1 and 23.12, 12.87, 0.12 and 1.09 exchangeable Ca2+, Mg2+, Na1+ and K1+, respectively, and had a pH value (in water) of 7.76 and EC value of 0.06 dS mat−1 the start of the experiment. The number of rhizobia nodulating peanut at Fedis soil was < 102 viable cell g−1 dry soil.

Sources of the Bradyrhizobium isolates

Rhizobial isolates used were recommended Bradyrhizobium sp. for Peanut (Arachis hypogeal L.). The bacteria used in this study were HUGNR-1, HUGNR-2, HUGNR-3, HUGNR-4, HUGNR-5, HUGNR-6, HUGNR-7 and HUGNR-8. These isolates were selected because they were previously found efficient isolates in N2 fixation under greenhouse conditions (Akuma 2010).

Sources of the cultivars

Recently released and high yielder cultivars of peanut (Werer-924 and Sedi) were obtained from lowland oil crop, Haramaya University, Ethiopia.

Preparation of inoculant

Purity was assured by routine plating on YEM agar supplemented with Congo red and white colony was selected for further use for inoculant production. White colony were transferred into 100 ml containing YEM (yeast extract mannitol) medium in 250 ml Erlenmeyer flasks and incubated at 28 °C on a shaker incubator at 150 rpm. After 48 h incubation, the number of viable cells in the culture broth was determined by the drop-plate method by counting colony-forming units (CFU) on YEMA plates (Somasegaran and Hoben 1994).

The carrier used for this experiment was decomposed filter cake after sterilized in autoclave at 121 °C. Filter cake based inoculant was prepared by mixing sterilized Filter cake and culture broth at 1:1 v w−1 under aseptic conditions and placed in an incubator at 25 °C for 1 week. After inoculation, the population of viable rhizobia cell was reached at 109 rhizobia g−1 of dry peat.

Experimental design and treatments

Ten treatments comprised of eight elite Bradyrhizobium, N fertilized 20 kg N ha−1) and an absolute control where no external inputs were applied were laid out as a randomized complete block design (RCBD) with three replications. The seeds were planted in 3 m rows with 10 cm spacing between plants and 40 cm between rows. The gross plot size was 3 × 3 m. Seeds which had been surface wetted, were mixed thoroughly with the peat culture and sown immediately. Triple-superphosphate (46 kg P2O5 ha−1) was uniformly applied for experimental plots. Inorganic N (20 kg N ha−1) as starter N was applied only for N-fertilized treatment. All inorganic fertilizer were incorporated in the soil before planting.

Indigenous rhizobial count

Undamaged and uniform seeds of peanut was sterilized using 0.2% HgCl2 and 95% ethanol (Vincent 1970) and then the germination was done in petri-dishes containing wetted filter paper. Three healthy seedlings were transferred to each of pot containing autoclaved sand (Vincent 1970) and thereafter thinned to one seedling per pot. From each soil sample, serial dilutions, ranging from 10−1 to 10−10 were prepared and added at a rate of 1 ml per pot. After 6 weeks, the numbers of nodulated plants were recorded and the most probable number (MPN) of rhizobia nodulating peanut was estimated (Brockwell 1963).

Agronomic data collection

At the late flowering stage at the R2 stage, five plants from internal rows were uprooted for nodulation parameters assessment. The soil adhering to roots and other plant parts was removed under running tap water. The nodules were collected and the dry weight of the plants and nodules were measured after drying the samples at 70 °C for 72 h. At harvest, plants obtained from 1 m2 were analyzed for yield determinations and the results expressed in kg ha−1.

Data analysis

Analysis of variance (ANOVA) was carried out using the randomized block design method. Least significance difference (LSD) was calculated to discriminate the effect treatment on the measured plant traits at 5% probability.

Nodulation

This field inoculation experiment was conducted at Fedis and Babile sites where the native rhizobia of peanut were 2.6 × 102 and 3.3 × 103 rhizobia of peanut g−1 soil, respectively as estimated by the MPN method. The result of the ANOVA showed that the main effect of inoculation (I), locations (L) and cultivar (C) and their two and three ways interaction significantly affected the pooled nodule number (NN) and nodule dry weight (NDW) at P < 0.05 (Table 1). However, the effect of cultivar on pooled NN and NDW across locations and L × C interaction effect on pooled NDW were not significant at P > 0.05, implying that the performance of the tested cultivar in terms of nodulation in these two locations are similar. This study showed that both cultivars performed equally in NDW production in both experimental locations. The result revealed that Bradyrhizobium inoculation did not improve significantly the pooled NDW compared with not inoculated.

Table 1

Analysis of variance of measured traits of peanut as affected by the main effect of Bradyrhizobium inoculation, location, and cultivar of peanut their two and three ways interaction

The main effect of location and C × I significantly influenced the % of effective nodules. This result suggests the presence of different effectiveness of native and inoculated Bradyrhizobium with the cultivar Werer-962 and Bati-Sedi. It was also found various effective nodules % at Babile and Fedis. Inoculating HUGNR-10 resulted in significantly increase the pooled NN over the uninoculated treatment (Table 2). However, inoculation did not affect the % of the effective nodule, indicating that the indigenous soil Bradyrhizobium sp. was able to induce effective nodules on peanut roots.

Table 2

Nodulation traits, shoot dry weight, number of seeds per pg and total pegs weight as affected by Bradyrhizobium inoculation, cultivar, and location

Treatments

NN

NDW

EffNN (%)

SDW

NSP

TPW

Consortium

138.75ab

0.2679ab

81.25a

51.7a

2.89ab

3384.3abc

HUGNR-10

158.83a

0.2703ab

83.25a

59.48a

2.72ab

3701.4a

HUGNR-11

121.83b

0.2332ab

83.25a

51.58a

2.72ab

3634.3a

HUGNR-12

137.58ab

0.2478ab

81.50a

53.97a

2.70ab

3752.3a

HUGNR-13

126.58b

0.2417ab

84.75a

55.03a

2.64ab

3493.1abc

HUGNR-18

139.17ab

0.2365ab

79.42a

54.81a

2.58ab

3613.4a

HUGNR-19

134.08ab

0.2160b

79.83a

50.35a

3.00a

3011.6c

HUGNR-29

133.00ab

0.2991a

85.17a

57.85a

2.92ab

3553.2ab

Uninoculated

115.75b

0.2394ab

79.33a

50.76a

2.45b

3062.5bc

N fertillized

137.08ab

0.2539ab

84.50a

43.84a

2.86ab

3518.5abc

F value

3.30**

2.28*

1.15ns

1.46ns

2.24*

4.64***

LSD (P < 0.05)

29.54

0.0715

9.66

16.73

0.52

539.46

Babile

118.883b

0.2314b

84.00a

57.38a

2.78a

3323.15b

Fedis

149.65a

0.2697a

80.45b

48.49b

2.72a

3621.76a

F value

57.46***

15.18***

7.15**

14.97***

0.71ns

16.23***

LSD (P < 0.05)

8.077

0.0196

2.64

4.575

0.1428

147.52

Werer 962

137.33a

0.2561a

82.43a

50.48b

2.24b

4019.44a

Bati-Sedi

131.20a

0.2450a

82.02a

55.39a

3.26a

2925.46b

F value

2.28ns

1.27ns

0.10ns

4.57*

201.02***

217.81***

LSD (P < 0.05)

8.08

0.0196

2.64

4.58

0.14

147.52

CV (%)

16.56

21.49

8.84

23.79

14.30

11.69

Means in the same column followed by the same letter are not significantly different at the 5% probability level by Tukey’s test

In general, the result revealed that significantly higher NN and NDW were found at Fedis than Babile site (Table 2). This could be associated the presence of high peanut rhizobia population and conducive environmental condition for nodulation at Fedis soil when compared to Babile soil. However, significantly higher % of effective nodules was recorded at Babile site than Fedis site. Both cultivars induced the statistically at par amount of NN, NDW and % of effective nodules.

All inoculated isolates including N fertilization except HUGNR-18 significantly increased the effective nodule % at Babile site (Fig. 1c). It was also found that only HUGNR-13 inoculation resulted in significant increase in effective nodule % at Fedis site. The present work also found that all isolates tested except HUGNR-18 inoculation caused the significant increased in effective nodules % with Bati-sedi cultivar (Fig. 2c). With Werer-962, a significant increase in effective nodules % was found due to HUGNR-11 and consortium inoculation and N fertilization.

The tested Bradyrhizobium showed different performance on NN and NDW production in both locations. At Babile site, except for HUGNR-11, all isolates including N fertilized plant gave the higher NN than the uninoculated treatment (Fig. 1a). At Fedis site, only HUGNR-11 inoculation resulted in increase in NN when compared to uninoculated treatment. This different result obtained at two locations may have arisen from higher native rhizobia population at Fedis than Babile site. Beside this, better NDW at Babile site were recorded due to Consortium isolates and HUGNR-29 inoculations but higher NDW than uninoculated at Fedis site due to all inoculation treatments, except HUGNR-10 and HUGNR-19 (Fig. 1b).

When the data stratified by cultivar, most of the tested Bradyrhizobium isolates except HUGNR-11, HUGNR-13 and HUGNR-29 improved the NN of the cultivar Werer-962 whilst all isolates inoculation with the cultivar Bati-Sedi cultivar resulted in higher NN than the control check (Fig. 2a). Only HUGNR-29 with werer-962 improved the NDW over the uninoculated treatment (Fig. 2b). The isolates which were resulted in a significantly increased NDW with Bati-Sedi were Consortium isolates, HUGNR-13, HUGNR-18 and HUGNR-29. This result indicates the permissiveness of the cultivar werer-962 for most of the tested isolates.

Shoot dry weight

The main effect of location and cultivar, and L × I and L × C on the pooled shoot biomass yield produced at late flowering stage across locations were significant at P < 0.05 (Table 1). Inoculation did not significantly (P > 0.05) increase the pooled shoot biomass of peanut (Table 2). Significantly higher pooled shoot biomass at the late flowing stage was recorded at Babile site than Fedis site, indicating Babile site could have a favorable condition for peanut as compared to the Fedis. The cultivar Bati-Sedi has recorded significantly higher pooled shoot biomass than the cultivar Werer-962.

Number of seeds per pod

The Bradyrhizobium inoculation and cultivar had significant effect on average number of seeds per pod across locations (Table 1). The lower number of seed per pod was measured in the uninoculated treatment. This indicates the need of N in the form of either inorganic or organic to improve the number of seeds per pod. A significantly higher average number of seeds per pod was recorded with the cultivar Bati-Sedi than the cultivar Werer-962. However, a number of seed per pod of peanut was not significantly differed which were obtained from two locations.

Total pods weight per hectare

The main effect of inoculation, cultivar and locations and their two ways interaction significantly varied the pooled total pods weight per hectare across locations (Table 1). Plants inoculated with HUGNR-10, HUGNR-11, HUGNR-12 and HUGNR-18 resulted in significantly increased total pod biomass (kg ha−1) as compared uninoculated treatment but did not differ statistically with N-fertilized treatment (Table 2). This implies that these isolate may provide a similar amount of N as it had been applied N in the N fertilized treatment. Inoculating HUGNR-12 increased the total pod’s weight by 22.5% over uninoculated treatment. Significantly higher average total pod weight was recorded at Fedis than Babile site. The cultivar werer-962 produced significantly higher average total pods weight than the cultivar Bati-Sedi.

100 seeds weight

ANOVA revealed that the main effect and their two and three interaction effect of three factors significantly influenced the pooled hundred seeds weight across locations (Table 1). However, none of the tested Bradyrhizobium isolates did improve significantly (P < 0.05) the pooled hundred seeds weight of peanut. Across the inoculation treatments, Babile site recorded significantly higher hundred seeds weight than Fedis site. The cultivar Werer-962 recorded significantly better average hundred seeds weight than the cultivar Bati-Sedi.

Total biomass yield

The ANOVA revealed that the main effects of inoculation, cultivar, and location and their two ways interaction effect were significant on the pooled total biomass yield per hectare of peanut across locations (Table 1). Inoculating HUGNR-29 significantly increased pooled total biomass yield by 26.2% compared with uninoculated but this biomass was statistically at par with N-fertilized treatment (Table 3). Both locations produced statistically at par amount of average total biomass yield of peanut. Significantly higher average total biomass yield across inoculation treatments was produced with the cultivar Werer 962 than Bati-Sedi.

Grain yield

The main effect of inoculation, cultivar and locations and their two and three ways interaction significantly affected the pooled grain yield of peanut across locations (Table 1). However, the main effect of location and L × C did not influence the pooled grain yield, showing that location different did not influence the grain yield production of the tested cultivars of peanut. All Bradyrhizobium inoculation including N fertilized treatments produced significantly higher pooled grain yield than the uninoculated treatment (Table 3). In both experimental locations, all Bradyrhizobium inoculation including N fertilization were found to increase the grain yield production (Figs. 1d, 2d). Moreover, all inoculation treatment including N application with both cultivars gave the higher grain yield than uninoculated.

Inoculating HUGNR-10 and HUGNR-29 plants were found to increase the pooled grain yield by 35.8 and 33.45% over the uninoculated treatment, respectively. When compared to the uninoculated, fertilization with N increased the grain yield by 34.5%. The difference in pooled grain yield production between two locations was not statistically significant at P > 0.05. The cultivar Werer-962 recorded significantly higher pooled grain yield than the cultivar Bati-Sedi.

Shelling %

Except for the main effect of inoculation, and three-way interaction, the remaining main and interaction effect on pooled shelling % of peanut was significant at P < 0.05 (Table 1). Inoculations and N application increased significantly the shelling percentage of a peanut compared with uninoculated (Table 3). Averaged across the inoculation treatments, significantly higher shelling % was recorded at Babile site than Fedis site. The cultivar Bati-Sedi was found to have significantly higher shelling % than Werer-962.

Total plant N and P concentration

The ANOVA revealed that none of the main and their interactions was affect significantly (P > 0.05) the total plant P concentration (Table 1). However, the main effect of location, cultivar and three ways interaction effect on pooled total plant N concentration was significant (P < 0.05). Inoculation had no significant effect on pooled plant N concentration, showing that indigenous and inoculated Bradyrhizobium could have been a similar efficacy in N2 fixation. Averaged total N plant concentration of peanut across the inoculation treatments at Fedis site was found to be significantly higher than that obtained from Babile site. The cultivar werer-962 accumulated significantly higher average plant tissue N than Bati-Sedi.

Use of inoculants in Ethiopia for food legume production including peanut is very infant. Due to low productivity of peanut in Ethiopia, enhancing the peanut-rhizobia symbiosis thereby improve the productivity using inoculation of effective rhizobia is essential. The native rhizobia population, the host genotypes and environmental condition are the major factors affecting the efficacy of inoculated rhizobia. The soil of the study sites had the native rhizobia capable of nodulating peanut greater than 102 g−1 of soil. Large number of native rhizobia in the two soils could be because of the presence of conducive soil properties (Yousef et al. 1987; Thrall et al. 2007; Drew et al. 2012) and cultivating the host plant for long time (Chemining’wa and Vessey 2006; Kimiti and Odee 2010). However, the previous experiment conducted this region showed that inoculation of locally isolated rhizobia improved the yield of common bean (Anteneh 2016), Fababean (Argaw et al. 2012a) and Field pea (Argaw et al. 2012b), though the soil had > 103 rhizobia g−1 soil. Therefore, this study was initiated to evaluate locally isolated Bradyrhizobium effectiveness on the productivity of peanut at major production areas of eastern Ethiopia.

Despite the natural rhizobia population has been large enough in the experimental soils, inoculation of selected isolated of native Bradyrhizobium significantly increased the pooled nodule number. However, the result revealed that inoculation did not affect significantly the pooled nodule dry weight and % of effective nodules of peanut Similarly, effective indigenous Bradyrhizobium isolate did not increase nodulation when the native rhizobia were abundant (Okogun and Sanginga 2003; Denton et al. 2009). On top of this, the present study found the non-significant effect of inoculation on pooled shoot dry weight, total plant N accumulation, and total P concentration. This result clearly indicated that the native rhizobia effective in supplying N to host plants as the inoculated isolates did. In line with this result, Kremer and Peterson (1983) found that the N2 fixation by peanut has been only increased when the inoculation improved the nodule weight.

When the data was stratified by location, at Fedis site, of tested isolates, consortium isolates and HUGNR-29 inoculating plants had significantly higher NDW than the uninoculated plants. At Babile site, except HUGNR-18, all tested isolates including N application increased significantly the NDW and % of the effective nodule. In line with this finding, inoculation of Rhizobium on promiscuous legume plants increased nodulation (Valladares et al. 2002). However, few numbers of tested rhizobia found effective at Fedis site could be associated with the presence of competitive and high rhizobia population nodulating peanut in the experimental soil. Similarly, Wielbo et al. (2012) highlighted that soil type and the host plant identity influenced the competitiveness of rhizobia. Competitive ability of rhizobia strains also determined by the metabolic potential in utilizing a diversity carbon and energy sources of tested isolates and the responsiveness of nod genes to flavonoid activation (Wielbo et al. 2007; Maj et al. 2010).

All isolates of Bradyrhizobium significantly increased the nodule number of the cultivar Werer-962 while a significant increase of this trait for the cultivar Bati-Sedi was found when inoculated with Consortium isolates, HUGNR-10, HUGNR-12, HUGNR-18 and HUGNR-19. Only HUGNR-29 inoculation increased the significantly nodule dry weight of the cultivar Werer-962. Effective nodules % of the cultivar Werer-962 significantly improved by isolate HUGNR-18 inoculation. The difference in strain survival, growth, and competition in the rhizosphere, the production of appropriate plant and bacterial signals might have been the causes of the difference in compatibility of tested isolated with tested cultivars of peanut (Wollum 1998). This observation was in agreement with the findings of Chen et al. (2003) that there has been peanut cultivars-Bradyrhizobium strain specificity in Argentina soil. With cultivar Bati-Sedi, most of the tested Bradyrhizobium increased the nodule dry weight and % of effective nodules. With regards to inorganic N application, the significant increased nodule dry weight and % of effective nodules found with Bati-Sedi but did not found with Werer-962. This indicates the variance effect of N application on nodulation of the two tested cultivars of peanut. This finding is confirmed by Abdel Wahab and Abd-Alla (1996) with soybean and showed that Soybean cv. Crawford showed a greater response to the low rate of N than cv. Clark.

The result of the present work has highlighted that inoculating HUGNR-29 was effective in significantly improving the pooled NDW and % of effective nodules. This isolate also improved the pooled total biomass and grain yield of peanut, indicating the need of inoculation for improving the effective nodule and production of peanut in the study sites as has been reported by Huang (1987). On top of this, all tested Bradyrhizobium inoculation including inorganic N application significantly increased the grain yield of peanut at Fedis and Babile sites. The positive effect of N application might be associated the presence of low concentration of nitrate in both soils which could act as an additional source of nitrogen until the plant start N2 fixation (Becana and Sprent 1987). However, the result showed that yield increases with inorganic N application was not statistically at par with HUGNR-29 inoculation, implying that Rhizobium inoculation can substitute the external inorganic N application. Elkan (1992) reported that the rhizobia-peanut symbiosis can derive N from the atmosphere as much as 152 kg N ha−1 which is 80% of the plant of the protoplasmic nitrogen. Moreover, about 55% of the plant N needs of peanut are delivered from the air via nitrogen fixation (Hardarson 1993).

The averaged nodule number and dry weight and total plant N accumulation across the inoculation treatments at Fedis were found to be significantly higher than those obtained from Babile. This difference could be because of the environmental conditions such as the effectiveness and competitiveness of the native rhizobia, soil condition and weather condition prevailed during the experimental time at both locations. Räsänen and Lindström (1999) found the different ability of isolate HAMBI2180 nodulating Acacia Senegal due to different soil temperature. The result the currect work found the non-significant difference in averaged % of effective nodules, total biomass and grain yield across the inoculation treatments which were recorded at Fedis and Babile. This indicates that indigenous rhizobia in Fedis and Babilale sites may have been similar effectiveness in N2 fixation and supplying N to the plant.

It was also found the non-significant difference in nodule number and dry weight and % of effective nodules recorded between the cultivar Werer-962 and Bati-Sedi. This finding is in line with the study on peanut cultivars (van Rossum et al. 1994) that had similar nodulation in neutral soil pH condition. It has been reported previously by Alwi et al. (1989) who tested 29 Bradyrhizobium against 16 cultivar of peanut and found that some of the isolates were effective with varieties and other showed a degree of specificity with cultivar. Chen et al. (2003) demonstrated that Bradyrhizobium displayed a varying degree of effectiveness, with some isolates being fairly effective with all tested cultivars and others with selected ones.

Despite the variation in nodulation, significantly higher averaged total pods weight, 100 seeds weight, total biomass and grain yield and shelling % across the inoculation treatments were found with Werer-962 than Bati-Sedi. These differences could have been due to inherent productivity potential difference between the two tested cultivars of peanut. Barbour et al. (1992) found that nodulation and N2 fixation by peanut varieties are found to be heritable and controlled by genetic make of the host plant. In the current work, the cultivar Werer-962 accumulated significantly higher plant N concentration than the cultivar Bati-Sedi. This N concentration difference between the cultivars could have been because of the fact that biomass production of the cultivar werer-962 was greater than the cultivar Bati-Sedi which cause variation in N2 fixation. Peoples et al. (2001) highlighted that 20–25 kg of shoot N has been fixed for every tone of legume shoot dry weight accumulated. Ankomah et al. (1996) found out that the good correlation between shoot N concentration and N2 fixation. In addition, the variation of N accumulation might also be related with inherent property of the tested peanut cultivars as has been reported by Nambiar and Dart (1980).

It is evident from the results of this study that inoculation of Bradyrhizobium is necessary in order to improve nodulation and seed yields of peanut. This result also revealed that the need of location and cultivar specific selection of Bradyrhizobium for better production of peanut in the study locations. It has been also found that N applied at starter rate requires to improve the yield of peanut. Further work is needed to determine whether combined application of N with Bradyrhizobium or/and their sole application can improve the peanut productivity in the study site. Moreover, multi-locational trials are required to check their effectiveness for the other agro-climatic conditions to give concrete conclusive to use the isolate as a candidate for national biofertilizer production.

Acknowledgements

This work was funded by the Ethiopian Institute Agricultural Research, Peanut Improvement Project. A special thanks to the field assistant for invaluable help with data collection and processing in both the field and the laboratory.

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The author declare no competing interests.

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